Today almost all polypeptides used in medicaments are prepared recombinantly. Due to strict regulatory guidelines and requirements, by-products have to be removed from the therapeutic polypeptide preparation as much as possible. Therefore, at least one chromatography step is employed in down stream processing of the bulk raw polypeptide after recombinant production. As the dimension of the chromatography equipment with respect to the yield of the fermentation process, especially the separation capacity of chromatography columns, is limited, a multitude of batches have to be processed in order to be able to provide the required amount of purified therapeutic polypeptide.
To ensure that each batch of the purified therapeutic polypeptide has the same pharmaceutical effect, a list of analytical parameters has to be fulfilled for each batch. This can only be achieved if the steps of the purification process operate consistently and efficiently. But, if one step of the purification process does not work properly the obtained product will most probably not pass the analytical tests and, in the worst case, this batch cannot be used. Therefore, it is necessary to provide methods for determining the performance and efficacy of purification steps.
Teeters, M. A. and Quinones-Garcia, I. (J. Chrom. A 1069 (2005) 53-64) report the evaluating and monitoring the packing behavior of process-scale chromatography columns by using the responses to conductivity-based pulse and step inputs derived from tracer experiments and in-process transitions, especially from measured residence time distributions. Norling, et al. (Norling, L., et al., J. Chrom. A 1069 (2005) 79-89) report the impact of multiple re-use of anion-exchange chromatography media on virus removal. The use of process data to assess chromatographic performance in production-scale protein purification columns is reported by Larson, et al. (Larson, T. M., et al., Biotechnol. Prog. 19 (2003) 485-492). Moscariello, J., et al., J. Chrom. A 908 (2001) 131-141 report the characterization of the performance of industrial-scale columns. The resolution and column efficiency in chromatography is reported by Vink, H., J. Chrom. 69 (1972) 237-242. Sarker, M. and Guiochon, G., J. Chrom. A 702 (1995) 27-44 report a study of the packing behavior of axial compression columns for preparative chromatography.
The use of an integrated form of the Gaussian distribution function allows for the description of the packed bed characteristics, whilst neglecting effects outside the packed bed itself, which potentially influence the evaluation (see e.g. PCT/EP2010/003813). The implementation of equipment characteristics in the evaluation of non-Gaussian distributions observed during the assessment of packed chromatographic beds has been described by Guiochon, G., et al. (FUNDAMENTALS OF PREPARATIVE AND NONLINEAR CHROMATOGRAPHY; Guiochon, G., et al. (eds), Elsevier Inc., San Diego (USA), 2nd edition (2006)).